This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. Elucidating mechanisms controlling metal accumulation and distribution in developing seed is critical for increasing the concentration of these micronutrients in crop seeds. Our research combines established genomic techniques, such as gene deletion and overexpression, with synchrotron x-ray fluorescence (SXRF) microspectroscopy and microtomography to characterize the function of genes involved in metal ion homeostasis in plants. SXRF was used to analyse dry mature Arabidopsis seeds and generated crucial information on metal localization and on the function of the tonoplastic Fe transporter VIT1. However, dry seed provides a limited view of metal homeostasis and does not reflect all the events occurring during seed development. SXRF analysis of developing seeds represents a necessary step to better understand how metal are accumulated and distributed in developing seeds. The proposed work at SSRL is divided in two main aims: first, we would like to image the metal distribution within seeds of wild-type Arabidopsis plants while they develop within the siliques. We will generate 2D elemental maps of entire siliques and 3D tomographic maps of individual seeds at the different key stages of development. Expected results will provide a detailed view of metal accumulation and metal transfer events during seed development. Second, we would like to characterize the function of Zn transporter in developing seeds, by examining alterations of metal distribution within developing seeds of loss-and-function mutants. Results will provide relevant information concerning the function of these Zn transporters in developing seeds, information that couldn?t be provided by alternative techniques. This experiment is even more crucial, because to date no genes have been implicated in Zn transport within developing seeds. The proposed work will provide a great deal of fundamental information that will contribute to our understanding of metal homeostasis in developing seeds.